520 PATHOGENIC MICROORGANISMS 



wall is prepared by shaving or plucking out the hair. A twenty-four- 

 hour pure culture on Loeffler's medium is emulsified in 20 c.c. of 

 normal salt solution and 0.15 c.c. of this suspension is injected intra- 

 cutaneously at a corresponding site into each of the two guinea-pigs. 

 One of these animals is given at the same time an intracardial injec- 

 tion of about 250 units of antitoxin, or is prepared by an intraperi- 

 toneal injection of antitoxin twenty-four hours before the tests are 

 made. Six cultures may be tested in this way on two animals. Viru- 

 lent strains produce a definitely circumscribed local infiltrated lesion, 

 which shows superficial necrosis in two to three days. In the control 

 pig the skin remains normal. This method, in the hands of Zingher, 

 gives results parallel to those obtained with the subcutaneous tests. 



Diphtheria Toxin. 1 Animals and man infected with B. diphtherise 

 show evidences of severe systemic disturbances and even organic de- 

 generations, while the microorganism itself can be found in the local 

 lesion only. This fact led even the earliest observers to suspect that, 

 in part at least, the harmful results of such an infection were attrib- 

 utable to a soluble and diffusible poison elaborated by the bacillus. The 

 actual existence of such a poison or toxin was definitely proved by 

 Roux and Yersin 2 in 1889. They demonstrated that broth cultures in 

 which B. diphtheria? had been grown for varying periods would remain 

 toxic for guinea-pigs after the organisms themselves had been removed 

 from the culture fluid by filtration through a Chamberland filter. 



METHODS OP PRODUCTION OP DIPHTHERIA TOXIN. While toxin can 

 be produced with almost all of the virulent diphtheria bacilli, there is 

 great variation in the speed and degree of production, dependent upon 

 the strain of organisms employed and upon the ingredients and reac- 

 tion of the medium upon which they are grown. Most laboratories 

 possess one or several strains of bacilli which are empirically known 

 to be especially potent in this respect. One of the most extensively 

 used, not only in this country but in Europe as well, is the strain 

 known as "Culture Americana," or " Park- Williams Bacillus No. 8," 

 an organism isolated by Dr. Anna Williams of the New York Depart- 

 ment of Health in 1894. Throughout more than ten years of cultiva- 

 tion this bacillus has retained its great power of toxin production. 



Because of the severity of cases of diphtheria in which the diph- 

 theria bacilli were associated with streptococci, many observers were 

 led to believe that the presence of streptococci tended to increase the 



, Cent. f. Bakt., 1887. 2 Roux and Yersin, loc. cit. 



